Our studies of the structure and function of opiate receptor subpopulations have continued using subpopulation specific irreversible inhibitors we developed earlier. Among these, BIT and FIT were shown to be mu and delta selective, respectively. In morphine tolerant rats selective upregulation of tritiated DADL binding sites was observed, and was confirmed by Scatchard analysis of tritiated DADL binding after FIT treatment of the membranes. Optimized conditions for tritiated bremazocine binding to kappa receptors have been defined in the rat and guinea pig brain. Treatment of rat and guinea pig brain sections with BIT and FIT followed by autoradiographic imaging with tritiated bremazocine revealed anatomical distribution of kappa sites. Optimized conditions to label mu, delta and kappa receptors have been utilized to determine receptor subtypes and density in the rat pituitary. Little if any mu and delta binding was observed, and the kappa binding was confined to the neural lobe where it was most dense in the external rim.